Chromatography is a well known separation technique whereby a number of analytes contained in a sample solution can be separated from one another. By using a suitable detector, the identity and amount of each analyte in the sample solution can be estimated.
One of the most useful forms of chromatography used for analytical purposes is gas chromatography, and particularly capillary gas chromatography. The sorbent material, or stationary phase, in this method can be a liquid or crosslinked polymer coated on the inner walls of the capillary column. An inert carrier gas such as helium is flowed through the column and a small amount of the sample solution is simultaneously vaporized and injected into the carrier gas ahead of the column. Once the sample is within the capillary column, the components of the sample are ideally separated into individual bands which move at different rates down the length of the column.
The mechanism by which this separation occurs is in general a continuous series of absorption and extraction steps. A component in the sample is absorbed into the stationary phase, and thereafter extracted into the carrier gas, or mobile phase, as it passes over the stationary phase. At any point in the column, the concentration of a component in the stationary phase is in equilibrium with the concentration of the component contained in the mobile phase. These two concentrations are related to one another by what is known as the partition coefficient (k), which is the concentration of a component in the stationary phase divided by the concentration of the component in the mobile phase. In this fashion, a moving band of the component is created, and since the partition coefficient for each component is ideally unique, each component contained in the sample will move at a different rate through the column.
The identity and amount of each component present in the sample can be determined by the detector, which is usually attached to a recording device which produces a plot (chromatogram) having separate peaks for each component in the sample. By comparing the length of time it takes for a component to pass through the column (the retention time) with standardized data for similar column conditions, one can estimate the identity of the component. The concentration of each component is normally determined by comparing the area beneath each component peak with the area beneath a standard peak of the component.
The partition coefficients are also influenced by the temperature of the column itself. Usually as the temperature increases, the partition coefficient correspondingly decreases, which in turn results in the component bands moving at a faster rate through the column. Due to this phenomenon, the temperature of the column can be tailored for the sample solution being analyzed. On the other hand, however, the column temperature suitable for resolving several components in a sample may not be a suitable temperature for resolving other components in the sample. At one temperature, for example, several components in a mixture, particularly the more volatile ones, may move through the column at a rate close to that of the carrier gas itself. The chromatogram peaks for such components will often overlap one another. At the same time, other components may move so slowly through the column that the analysis period is too long and/or the component bands spread or diffuse to an unacceptable width.
One means for addressing these problems is to use temperature programming, whereby the temperature of the entire column is increased in a controlled manner while the sample solution is passing through the column. By this method, the faster moving component bands can often be resolved from one another at a relatively low column temperature and then the component bands which typically move at a slower rate through the column are speeded up by increasing the column temperature. Temperature programming is usually accomplished by placing the entire column inside an oven whose temperature can be controlled.
Even with temperature programming, however, some sample mixtures cannot be properly analyzed due to overlapping component peaks, band dispersion, or a combination of both. In addition, another drawback of temperature programming is that the oven must be recycled back to its initial temperature before each subsequent sample can be run through the column, thereby limiting the number of samples that can be run in a given period.
An alternative arrangement that has been discussed in the literature is the use of temperature gradients, or chromathermography, as some have referred to it. In this approach, the temperature within the column varies along its length from a relatively high temperature at the inlet of the column to a relatively lower temperature at the outlet of the column, i.e., a negative temperature gradient. An additional modification of this method, is where the column temperature also varies with time, thereby producing a tim-variable temperature gradient along the length of the column, e.g., a beginning gradient of from 150.degree. C. at the column inlet to 50.degree. C. at the column outlet and a final gradient of from 250.degree. C. at the column inlet to 100.degree. C. at the column outlet. While both of these approaches have been discussed in the literature, heretofore there has not been an accepted way of generally applying these techniques to capillary columns.
By applying a negative temperature gradient along the length of the column all of the components can be moved through the column more quickly, while at the same time the resolution between chromatogram peaks is often improved, and band spreading or diffusion is often decreased. Even better results can be obtained when the temperature gradient varies with time.
While time-variant temperature gradient chromatography has previously been attempted, the methods and devices used to achieve this effect were not accepted for commercial use. Nerheim, in an article entitled "Gas-Liquid Chromathermography," described the use of a motor-driven heater fitted around a gas chromatography column. This device, and similar ones used by others, however, are bulky for everyday use and are not easily applied to long capillary columns (which can often be 60 meters in length or longer). Moreover, these devices are generally incapable of producing a linear temperature gradient in the column.
Another method previously used to establish a time-variable thermal gradient in a column involves painting the outside of the column with varying levels of resistive paint along the length of the column. By applying electric current to the outer layer of the column, a temperature gradient is produced. This thermal gradient can be varied with time by changing the level of current provided to the resistive paint. One problem with this technique, however, is that each column must be separately painted.
A summary of other impractical methods for establishing temperature gradients in capillary columns which have been tried is contained in the article by Berezkin, et al. entitled "Temperature Gradients in Gas Chromatography." FIG. 12 of this article, for example, shows one particular method for applying a time-variable temperature gradient along a chromatography column. The device shown in FIG. 12, however, suffers from the drawback of not easily being capable of producing a linear temperature gradient.
Consequently, heretofore, there has not been available any commercially accepted method and apparatus for establishing a linear, time-variable temperature gradient in a capillary chromatography column.